Project Summary/Abstract The Vascular Contributions to Cognitive Impairment and Dementia (VCID) are recognized and undeniable. In addition to Cerebral Amyloid Angiopathy (CAA), present in more than 90% of individuals with Alzheimer's disease and in many elderly people, chronic cardiovascular disease is also known to impair brain microvascular function, leading to cognitive decline. However, the interactive nature of CAA and chronic cardiovascular challenges and the mechanisms through which they contribute to cerebrovascular dysfunction are poorly understood. Our work has identified molecular pathways responsible for A?-mediated microvascular endothelial dysfunction. We found that A? induces overexpression/activation of endothelial TRAIL (TNF- related apoptosis inducing ligand) death receptors (DR) DR4 and DR5, new targets with enormous impact for CNS as well as vascular stress/death. DRs trigger mitochondrial dysfunction, with release of pro-apoptotic factors and reactive oxygen species. Our interventions targeting these pathways have reversed the pathologic effects of vascular A? in vitro and in vivo. Intriguingly, chronic cardiovascular conditions associated with cerebrovascular pathology are known to contribute to similar endothelial stress pathways, and may further impact these mechanisms. We hypothesize that chronic cardiovascular challenges, such as cerebral hypoperfusion, hypertension and hyperhomocysteinemia (HHC), synergize with cerebrovascular A? to enhance TRAIL DR activation and mitochondrial dysfunction in cerebral endothelial cells, promoting neurovascular failure in Alzheimer's disease. In Aim 1, using human cerebral microvascular endothelial cells in culture and human bio-fluids, we will test the hypothesis that chronic cardiovascular challenges synergistically increase the effects of A? on DR-mediated cell stress/death, mitochondrial dysfunction, and blood brain barrier (BBB) permeability. We will use in vitro cellular, molecular and imaging approaches (Aim 1A), as well as high sensitivity fluid biomarker discovery in elderly human subjects (Aim 1B). In Aim 2, we will test the hypothesis that chronic cardiovascular insults (hypertension and HHC) synergistically affect DR and mitochondrial endothelial dysfunction in a mouse model of cerebrovascular amyloidosis (TgSwDI), contributing to microvascular and cognitive impairment. We will utilize molecular, in vivo imaging and behavioral approaches at early and established disease stages. In Aim 3 we will apply new and repurposed interventions (DR silencing and FDA-approved carbonic anhydrase inhibitors) to test the hypothesis that decreasing DR activation and mitochondrial dysfunction will prevent endothelial stress and BBB damage induced by the combination of A? and chronic cardiovascular challenges in cells and mice. If successful, this study will reveal modifiable molecular mechanisms underlying mixed cerebrovascular disease, as well as biomarkers for cerebrovascular damage in Alzheimer's patients based on a more detailed understanding of molecular pathways, with potentially rapid impact on clinical practice and public health.